This invention deals with therapeutic and prophylactic treatment of age-related problems with humans. More specifically, a new use for existing medicaments that will counteract the aging process on a cellular levelxe2x80x94particularly in the brainxe2x80x94is disclosed.
Normal aging in humans is recognized as producing some or all of the following typical physiological results:
1. Brain weight is reduced by 15%
2. Blood flow to the brain is reduced by 20%
3. Body water content is reduced by 18%
4. Body weight is reduced by 12%
5. Nerve conduction velocity is reduced by 10%
6. Number of nerve fibers in nerves are reduced by 37%
7. Decreased amounts of enzymes and coenzymes
8. Decreased amounts of neurotransmitters
9. Depletion of oxidative, phospohorelative enzymes
10. Apoptosisxe2x80x94chronic neuronal atrophy
In describing their work in an article entitled xe2x80x9cStudies on Age-Dependent Ozonide Changes in Human Cerebral Cortex,xe2x80x9d (by Reichlmeier K., Ermini M., and Schlecht H. P.xe2x80x94Aktuelle Gerontol 1978 August, 8(8):44-8), the authors report that they investigated the activity of various enzymes of human brains obtained at autopsy and covering an age range from 19 to 91 years. Protein kinase, which mediates the information carried by the second messenger, cyclic AMP (3xe2x80x2,5xe2x80x2-cyclic adenosine monophosphate), does not show age-related changes of basal activity. Cyclic AMP-dependent activation of protein kinase remains nearly constant up to 60 years of life, but it undergoes a distinct and progressive decline between 60 and 90 years. In the corpus striatum, no age related changes of cyclic AMP-dependent protein kinase activity were observed. The activity of carbonic anhydrase exhibits, in both human cortex and corpus striatum, an age-dependent decrease that also begins after the sixth decade of life. These neurochemical changes may well be related to morphological and physiological changes occurring in the aging brain. They begin after the 60th year of life.
The following represents an essential chemical reaction that takes place in human tissue:                                           Glucose            +            Oxygen                                (                          WITHIN              ⁢                              xe2x80x83                            ⁢              CELLS                        )                          →                                                                              H                  2                                ⁢                                  xe2x80x83                                ⁢                O                            +                              C                ⁢                                  xe2x80x83                                ⁢                                  O                  2                                                      =                                          H                *                            +                              HCO                3                -                                                                        Carbonic              ⁢                              xe2x80x83                            ⁢              Anhydrase              ⁢                              xe2x80x83                            ⁢              Enzyme              ⁢                              xe2x80x83                            ⁢              present                                      (                              reversible                ⁢                                  xe2x80x83                                ⁢                reaction                            )                                                          Eq        .                  xe2x80x83                ⁢                  (          1          )                    
Glucose is irreversibly oxidized within the cells to produce water and carbon dioxide. In the presence of a catalyst, especially a carbonic anhydrase enzyme (of which several forms exist, of which the form present depends upon the type of tissue cells present), the water and carbon dioxide reversibly produce a hydrogen ion and a bicarbonate ion.
Carbonic anhydrase is a zinc-containing enzyme-that catalyzes-the reversible CO2 hydration reaction illustrated in Eq. 1. The mitochondria of cells of different tissues and organs produces different specific carbonic anhydrase enzymes that maintain the equilibrium of the above equation in all spacesxe2x80x94cellular, interstitial, and vascularxe2x80x94as illustrated in FIG. 1. At least seven carbonic anhydrase variants, called xe2x80x9cisozymesxe2x80x9d have been identified. The literature may refer to these as xe2x80x9ccarbonic anhydrases I through VIIxe2x80x9d or xe2x80x9cCAS I-VIIxe2x80x9d. We here refer to this selectivity as xe2x80x9ccell-specificxe2x80x9d and the particular carbonic anhydrase isozyme present as being a xe2x80x9ccell-specific carbonic anhydrase enzyme.xe2x80x9d
Hydrogen ion produced by carbonic anhydrase enzymes is acted upon by cytochrome system, which is then utilized as the energy source of the ion pump that maintains the integrity of the cell membrane comprising and enclosing each cell. It is also thought to be a source of the brain""s electric current. This process is schematically illustrated in FIG. 1, presented here with no further discussion.
Disruption of the process that includes Eq. 1 causes depolarization of the cell wall membrane, hence sodium (Na), water (H2O), and other chemicals can enter the cell in uncontrolled amounts and potassium (K) can exit uncontrollably, leading to the death and destruction of the involved cells; cellular edema follows. As this edema progresses, the cell dies. Along with the progressive and gradual death of cells, gliosis followsxe2x80x94hence aging in the brain occurs.
In aging, there has been observed a progressive decrease in levels of enzymes of which carbonic anhydrase enzyme is one. Authors W. Meier-Ruge, P. Iwangoff, K. Reichlmeier, and P. Sandoz, in xe2x80x9cNeurochemical findings in the Aging Brain (Adv. Biochem Psychopharmacology 1980;23;323-38) include carbonic anhydrase in their studies of normal aging on enzymes in the human brain cortex and putamen. Their study shows carbonic anhydrase, which they cites as being important to the regulation of the pO2/pCO2 ratio in the brain tissue, demonstrates a significant decline with increasing age. Thus, pCO2-dependent regulation of tissue pH, ionic transport processes, and cerebral blood flow regulation have the tendency to become more and more unstable, they observe.
Authors E. Cabisco and R. L. Levine, in xe2x80x9cCarbonic anhydrase III. Oxidative modification in vivo and loss of phosphatase activity during agingxe2x80x9d (J. Biol. Chem. 1995 June 16;270(24): 14742-7), describe their utilizing an immunochemical method for detection of oxidatively-modified proteins, through which method they identified a protein in rat liver that was highly oxidized. It was purified to homogeneity and identified as carbonic anhydrase isozyme III. Its characteristics match those previously described for protein that was lost during aging of the rat, senescence marker protein-1. In their experiments, carbonic anhydrase III was purified from rats aged 2, 10, and 18 months and the proteins were characterized. All three preparations were highly oxidative modified, as assessed by their carbonyl content. The enzyme (carbonic anhydrase III) has three known catalytic activities, and the specific activities for carbon dioxide hydration and for ester hydrolysis decreased during aging by approximately 30%. However, the third activity, that of a phosphatase, was virtually lost during aging. While the physiologic role of carbonic anhydrase III is unknown, these authors suggest that it functions as an oxidizing environment, which leads to its own oxidative modification.
Carbonic anhydrase enzyme has been used to augment the extracellular pH buffering in the cerebral cortex of rats (Journal of Neurophysiology 1995 October ""74(4):1806-9). It is known that the blood-brain barrier in animals is incomplete compared to that of humans where the blood-brain barrier is complete and a formidable barrier to chemical transport. Substances that prove efficacious in affecting the brain chemistry of animals are not necessarily efficacious in the brains of human beings because they cannot pass through the more complete blood-brain barrier in humans. Carbonic anhydrase enzymes appear to traverse the blood-brain barrier in humans. Although some researchers equivocate on this concept, most of the medical community accepts the idea that carbonic anhydrease enzymes traverse the blood-brain barrier in humans as fact, especially regarding the carbonic anhydrase enzyme referred to as CA-II.
As far as can be determined from the literature, cell-specific carbonic anhydrase enzymes have never been used to restore to a higher level the carbonic anhydrase enzymes that are lacking due to decreased levels due to normal aging. At least some of the carbonic anhydrase isozymes have been extracted from animal tissue, isolated, and studied for molecular structure. This shows that the enzymes can be isolated and made available for administration to a patient for therapeutic or prophylactic-treatment.
In U.S. Pat No. 5,972,684, Bandman et al. tell us:
xe2x80x9cEight enzymatic and evolutionarily related forms of carbonic anhydrase are currently known to exist in humans: three cytosolic isozymes (CAI, CAII, and CAIII, two membrane-bound forms (CAIV and CAVII), a mitochondrial form (CAV), a secreted salivary form (CAVI) and a yet uncharacterized isozyme. Isoforms show a characteristic motif. (See, e.g., http//expasy.hcuge.ch). Though the isoenzymes CAI, CAII, and bovine CAIII have similar secondary structure and polypeptide-chain fold, CAI has 6 tryptophans, CAII has 7 and CAIII has 8 (Boren, K. et al. (1996) Protein Sci. 5(12):2479-2484). CAII is the predominant CA isoenzyme in the brain of mammals.xe2x80x9d
xe2x80x9cInhibition and activation of CA provide information about CA stricture and activity. Vasodilating prostaglandins E1, E2 and I2 inhibit CA in vitro and in vivo and may inhibit the involvement of CA in gastric acid secretion. Nonsteroidal anti-inflammattory drugs which reduce the activity of cyclooxygenase and prostaglandin production have also been observed to activate CAI and CAII in a dose-dependent noncompetitive manner. The pre-prostaglandin cyclooxygenase appears to maintain an inverse relationship with CA, probably mediated by the pH variations associated with carbonic anhydrase activity (Puscas, I. (1996) J. Pharmacol. Exp. Ther. 277(3):1464-1466). Both prostaglandins E2 and I2 inhibit gastric acid output. Prostaglandin. E2 inhibits egress of norepinephrine from sympathetic nerve terminals.xe2x80x9d
The Bandman et al. patent teaches another carbonic anhydrase, CA-VIII, the subject of their patent. The present patent does not deal with nor address CA-VIII.
Patients having a carbonic anhydrase VI (CA-VI) deficiency have been treated with orally-administered zinc in an effort to stimulate the synthesis/secretion of CA-VI and the successful results were reported in the American Journal of Medical Science (Efficacy of exogenous oral zinc in treatment of patients with carbonic anhydrase VI deficiency, by Henkin, R. I., Martin, B. M., and Agarwal, R. P.xe2x80x94Am J Med Sci 1999 December;318(6):392-405). Thus, it is shown-that the synthesis/secretion of carbonic anhydrase can, indeed, be stimulated by compounds administered orally.